The objectives of the proposed research are to determine how microvilli of intestinal epithelial cells move and to determine the molecular basis for actin filament-membrane attachment using the brush border as a model system. I will characterize microvillar motility through studies on brush border motility in vivo and in vitro using motile models. The possible link between Ca ions transport and microvillar movement will be determined through studies on the motility and contractile proteins of brush borders isolated from intestinal epithelial cells of vitamin-D deficient animals. The basis for Ca ions regulation of motility will be investigated. I will determine if phosphorylation of brush border myosin is a requirement for the Ca ions induced contraction of microvilli observed in in vitro models. The presence of troponin-like regulatory proteins will also be investigated. I will extend previous observations on the characterization and localization of brush burder contractile proteins. Proposed experiments include: (a) isolation and characterization of microvilli, (b) characterization of a cytoskeleton associated CaMg ATPase, (c) characterization of brush border alpha actinin and (d) immunoelectron microscopic localization of brush border contractile proteins. I will identify possible linker molecules in the attachment of actin filaments to the brush border membrane using photoactivated membrane probes. The organization of the brush border membrane during microvillus growth will be determined through freeze fracture studies on the hydrostatic pressure induced dis-assembly and reassembly of microvilli.